Proliferation of normal cells is strictly controlled by the ordered repetition of the cell cycle consisting of interphase (G1 phase)-DNA synthesis phase (S phase)-interphase (G2 phase)-mitotic phase (M phase). In cancer cells, the cell cycle is out of control and abnormal cell proliferation is induced. One of the most important cell cycle regulators is Rb protein, which is known as a tumor suppressor gene product. Rb protein directly binds to the E2F/DP complex, a transcription factor essential for the initiation of S phase, and thereby inhibits its activity. Rb protein is phosphorylated and inactivated by multiple cyclin-dependent kinase (CDK) complexes in late G1 phase, thus promoting the transcription of E2F/DP to initiate S phase. The activity/expression of the CDK complexes which phosphorylate Rb protein are controlled through negative regulation by CDK inhibitor proteins such as p27, p21 and p16. Such an Rb protein-based cell cycle control mechanism is referred to as RB pathway. This cell cycle control mechanism, along with the p53 pathway, plays the most important role in controlling malignant transformation of cells.
p27 protein is one of the seven cyclin-dependent kinase (CDK) inhibitor proteins, which are known as negative regulators of the cell cycle. p27 protein binds to cyclin D/CDK4, cyclin E/CDK2, etc. and thereby inhibits their activities. Reduced expression of p27 protein allows kinases such as cyclin D/CDK4 and cyclin E/CDK2 to phosphorylate and inactivate the tumor suppressor gene Rb, thus inducing transcriptional activation of E2F/DP. E2F/DP transcribes genes necessary for cell cycle progression into DNA synthesis phase (S phase), thereby promoting cell proliferation. It has been known that forced expression of p27 protein in cancer cells arrests the cell cycle at G1 phase, thereby inhibiting cell proliferation.
Mutation or deletion of the p27 gene is rarely observed in cancer cells. In many cancers, however, abnormal subcellular localization or enhanced expression of its regulators such as Skp2 reduces the expression of p27 protein in nuclei, thereby promoting cell proliferation. The human p27 gene promoter was cloned (Non-patent document 35) and has been used as a useful research tool to attempt to elucidate the control mechanism of the p27 gene in cancer cells. As of now, the involvement of NF-Y in the p27 promoter activation mechanism has been reported. Specifically, it has been reported that NF-Y binds to the CCAAT box in the p27 promoter, thereby promoting p27 transcription (Non-patent document 36). Also, it has been reported that in leukemia cell lines, NF-Y and Sp1 inhibit cell proliferation and promote cell differentiation through a response of vitamin D3 (Non-patent document 37). Clinically, reduced expression of p27 protein is a reliable prognostic marker that correlates with poor prognosis of cancers, such as breast cancer, colon cancer, non-small cell lung cancer and prostate cancer, and it is suggested that reduced expression of p27 protein is associated with cancer cell proliferation. Further, in other cell proliferative disorders (rheumatism, inflammation, etc.), cyclin-dependent kinase activation associated with reduced expression of p27 protein results in abnormal cell proliferation, which is considered one of the causes of the disorder. For example, it has been shown that when p21 or p16, which is also a CDK inhibitor protein like p27, is transgenically expressed in animal models of arthritis, the exacerbation of rheumatism is suppressed (Non-patent documents 38 to 40). Also, it has been shown that an agent capable of increasing the expression of p27 protein may have therapeutic potential in the treatment of rheumatism. Thus, an agent capable of inducing p27 protein and increasing its expression is expected to be useful in the treatment of various diseases associated with cell proliferation, such as cancer, rheumatism and inflammation.
Meanwhile, it has already been revealed that coumarin derivatives, i.e., compounds which have a coumarin skeleton as a core structure, and in which the skeleton is derivatized at various positions, have different pharmacological effects depending on the position at which a chemical modification occurs (Non-patent document 31). For example, warfarin, which has antithrombogenic activity, is well known as a drug having a coumarin skeleton (Non-patent document 1). Furthermore, coumarin derivatives which exert antitumor activity by acting on various different target proteins, or coumarin derivatives which inhibit proteins associated with antitumor activity have been obtained by chemical modification at different positions of the core structure.
Coumarin derivatives that exert antitumor activity by inhibiting steroid sulfatase have been reported (Non-patent documents 2 to 6). Among them, compounds which are currently in clinical testing form a cycloalkyl group at the 3- and 4-positions of the coumarin skeleton, and has a sulfamate group at the 7-position. Their application to breast cancer is being considered from the viewpoint of pharmacological action.
Furthermore, as for coumarin derivatives which exert antitumor activity through binding to the estrogen receptor, a group of compounds having a characteristic substituent at the 4-position have been reported. Specifically, there have been reported a group of compounds having an arylalkyl group at the 4-position and having substituents at the 3- and 7-positions (Patent document 1), and a group of compounds in which a phenyl group is directly bound to the skeleton at the 4-position, and which have a phenoxy group at the 3-position (Patent document 2).
Also, as for a coumarin derivative exhibiting Raf inhibitory activity and exhibiting antitumor activity in cells, a compound having a 6-pyrazinyloxy group at the 7-position has been reported (Patent document 5).
In addition, known are several coumarin derivatives whose target proteins are unknown, and which are reported to exhibit antitumor activity. These include a group of compounds derived from natural sources (Non-patent documents 7 to 12) and a group of new compounds obtained by chemical synthesis (Non-patent documents 13 to 23, 32 to 34). As for the new compounds obtained by chemical synthesis, there have been reported, for example: a compound having alkoxy groups at the 5-, 6- and 7-positions of the coumarin skeleton (Non-patent document 13); a compound having an alkoxy group only at the 7-position of the coumarin skeleton (Non-patent document 14); a compound having an enone functional group at the 6- or 7-position of the coumarin skeleton (Non-patent document 15); a compound having a methyl group at the 4-position of the coumarin skeleton and substituents at the 7- and 8-positions (Non-patent document 16); a compound having substituents at all of the 4-, 5-, 6-, 7- and 8-positions of the coumarin skeleton (Non-patent document 17); a compound in which an amide group, ester group or sulfonamide group is directly bound to the coumarin skeleton at the 3-position, and which has a substituent at the 6- or 8-position (Non-patent documents 18 and 19); a compound in which an amide group is directly bound to the coumarin skeleton at the 3-position, and which has a substituent at the 7-position (Patent document 3 and Non-patent document 20); a compound having substituents at the 6- and 7-positions of the coumarin skeleton (Non-patent document 21); a compound having a hydroxy group at the 7-position of the coumarin skeleton and a nitro group at an appropriate position of the 3-, 6- and 8-positions (Non-patent documents 22 and 23); and a compound having a methoxy or hydroxy group at the 7-position of the coumarin skeleton, a phenyl group at the 3-position and a substituent at the 4-position.
There have also been reported: a compound having a substituent with a nitrogen atom (diethylamino group etc.) at the 7-position, a cyano group at the 4-position and a heteroaryl group at the 3-position (optionally with no substituent at the 4-position) (Non-patent document 32); and a compound having a heteroaryl group at the 3-position and a methyl group, halogen atom, nitro group, etc. at the 6-, 7- or 8-position (Non-patent document 33). There have also been reported examples of using a compound with a coumarin structure as a ligand for a Pd compound having antitumor activity in cells (Non-patent document 34).
As for compounds which exhibit target protein inhibitory activity, and which are likely to have antitumor activity despite the absence of a report dealing with their antitumor activity, there have been reported coumarin derivatives exhibiting TNFα inhibitory activity (Patent document 4 and Non-patent documents 24 to 28), aromatase inhibitory activity (Non-patent document 29), MEK inhibitory activity (Non-patent document 30), or the like. In compounds mentioned in these reports, the substituents are located at the 3-, 4-, 6- or 7-position of the coumarin skeleton.
As described above, although several coumarin derivatives with antitumor activity are known, few of the compounds exhibit a sufficiently high antitumor activity to put it to practical use as an anticancer drug. Also, no coumarin derivatives with p27 protein-inducing activity are known. Therefore, compounds which have practical p27 protein-inducing activity and can be used as an active ingredient of an antitumor agent etc. are still strongly sought.    Patent document 1: International Publication WO 2000/039120    Patent document 2: International Publication WO 2004/069820    Patent document 3: International Publication WO 2003/024950    Patent document 4: International Publication WO 2002/008217    Patent document 5: International Publication WO 2006/067466    Non-patent document 1: Ansell, J.; Bergqvist, D; Drugs 2004, 64, 1-5    Non-patent document 2: Purohit, A.; Woo, L. W. L.; Chander, S. K.; Newman, S. P.; Ireson, C.; Ho, Y.; Grasso, A.; Leese, M. P.; Potter, B. V. L.; Reed, M. J.; J. Steroid Biochem. Mol. Biol. 2003, 86, 423-432    Non-patent document 3: Lloyd, M. D.; Pederick, R. L.; Natesh, R.; Woo, L. W. L.; Purohit, A.; Reed, M. J.; Acharya, K. R.; Potter, B. V. L.; Biochem. J. 2005, 385, 715-720    Non-patent document 4: Purohit, A.; Woo, L. W. L.; Potter, B. V. L.; Reed, M. J.; Cancer Research 2000, 60, 3394-3396    Non-patent document 5: Woo, L. W. L.; Howarth, N. M.; Purohit, A.; Hejaz, A. M.; Reed, M. J.; Potter, B. V. L.; J. Med. Chem. 1998, 41, 1068-1083    Non-patent document 6: Woo, L. W. L.; Purohit, A.; Reed, M. J.; Potter, B. V. L.; J. Med. Chem. 1996, 39, 1349-1351    Non-patent document 7: Lopez-Perez, J. L.; Olmedo, D. A.; Olmo, E. D.; Vasquez, Y.; Solis, P. N.; Gupta, M. P.; Feliciano, A. S.; J. Nat. Prod. 2005, 68, 369-373    Non-patent document 8: Ito, C.; Itoigawa, M.; Mishina, Y.; Filho, V. C.; Enjo, F.; Tokuda, H.; Nishino, H.; Furukawa, H.; J. Nat. Prod. 2003, 66, 368-371    Non-patent document 9: Chen, Y-C.; Cheng, M-J.; Lee, S-J.; Dixit, A-K.; Ishikawa, T.; Tsai, I-L.; Chen, I-S.; Helv. Chim. Acta 2004, 87, 2805-2811    Non-patent document 10: Lee, K-H.; Chai, H-B.; Tamez, P. A.; Pezzuto, J. M.; Cordell, G. A.; Win, K. K.; Tin-Wa, M.; Phytochemistry 2003, 64, 535-541    Non-patent document 11: Chaturvedula, V. S. P.; Schilling, J. K.; Kingston, D. G. I.; J. Nat. Prod. 2002, 65, 965-972    Non-patent document 12: Madari, H.; Panda, D.; Wilson, L.; Jacobs, R. S.; Cancer Research 2003, 63, 1214-1220    Non-patent document 13: Riveiro, M. E.; Shayo, C.; Monczor, F.; Fernandez, N.; Baldi, A.; De Kimpe, N.; Rossi, J.; Debenedetti, S.; Davio, C.; Cancer Letters 2004, 210, 179-188    Non-patent document 14: Baba, M.; Jin, Y.; Mizuno, A.; Suzuki, H.; Okada, Y.; Takasuka, N.; Tokuda, H.; Nishino, H.; Okuyama, T.; Biol. Pharm. Bull. 2002, 25, 244-246    Non-patent document 15: Chen, Y-L.; Wang, T-C.; Tzeng, C-C.; Helv. 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